Stage of infection with Mycobacterium avium subsp. paratuberculosis impacts expression of Rab5, Rab7, and CYP27B1 in macrophages within the ileum of naturally infected cows

Taylor L T Wherry; Mark Heggen; Adrienne L Shircliff; Shankumar Mooyottu; Judith R Stabel

doi:10.3389/fvets.2023.1117591

Stage of infection with Mycobacterium avium subsp. paratuberculosis impacts expression of Rab5, Rab7, and CYP27B1 in macrophages within the ileum of naturally infected cows

Front Vet Sci. 2023 Feb 3:10:1117591. doi: 10.3389/fvets.2023.1117591. eCollection 2023.

Authors

Taylor L T Wherry^{1

2}, Mark Heggen³, Adrienne L Shircliff², Shankumar Mooyottu¹, Judith R Stabel²

Affiliations

¹ Department of Veterinary Pathology, College of Veterinary Medicine, Iowa State University, Ames, IA, United States.
² United States Department of Agriculture-Agricultural Research Service (USDA-ARS), National Animal Disease Center, Ames, IA, United States.
³ Department of Nutritional Sciences, College of Agricultural and Life Sciences, University of Wisconsin-Madison, Madison, WI, United States.

Abstract

Introduction: Macrophages are the preferential target of Mycobacterium avium subsp. paratuberculosis (MAP), the etiologic agent of ruminant paratuberculosis. Uptake of pathogens by intestinal macrophages results in their trafficking through endosomal compartments, ultimately leading to fusion with an acidic lysosome to destroy the pathogen. MAP possesses virulence factors which disrupt these endosomal pathways. Additionally, levels of serum vitamin D₃ have proven relevant to host immunity. Dynamics of endosomal trafficking and vitamin D₃ metabolism have been largely unexplored in bovine paratuberculosis.

Methods: This study aimed to characterize expression of early and late endosomal markers Rab5 and Rab7, respectively, within CD68+ macrophages in frozen mid-ileum sections harvested from cows at different stages of natural paratuberculosis infection. Additionally, factors of vitamin D₃ signaling and metabolism were characterized through expression of vitamin D₃ activating enzyme 1α-hydroxylase (CYP27B1), vitamin D₃ inactivating enzyme 24-hydroxylase (CYP24A1), and vitamin D₃ receptor (VDR) within CD68+ ileal macrophages.

Results and discussion: Cows with clinical paratuberculosis had significantly greater macrophage and MAP burden overall, as well as intracellular MAP. Total expression of Rab5 within macrophages was reduced in clinical cows; however, Rab5 and MAP colocalization was significantly greater in this group. Intracellular Rab7 colocalization with MAP was not detected in subclinical or Johne's Disease negative (JD-) control cows but was present in clinical cows. Additionally, macrophage CYP27B1 expression was significantly reduced in clinical cows. Taken together, the results from this study show disparate patterns of expression for key mediators in intracellular MAP trafficking and vitamin D metabolism for cows at different stages of paratuberculosis.

Keywords: CYP24A1; CYP27B1; Mycobacterium avium subsp. paratuberculosis; Rab5; Rab7; bovine; macrophage; vitamin D.

Grants and funding

This study was funded through USDA-ARS CRIS Project 5030-32000-235.